Food Science and Preservation (한국식품저장유통학회지)

The Korean Society of Food Preservation (한국식품저장유통학회)
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Inhibition Effect of Achyranthes japonica N. Root Extract on Cathepsin B

우슬뿌리 추출물의 Cathepsin B에 대한 저해효과

  • Lee Ka-Soon (Chungnam Agricultural Research and Extension Services) ;
  • Lee Jin-Il (Chungnam Agricultural Research and Extension Services) ;
  • Lee Jong-Kuk (Chungnam Agricultural Research and Extension Services) ;
  • Lee Jeong (Chungnam Agricultural Research and Extension Services) ;
  • Kim Gi-Don (Chungnam Agricultural Research and Extension Services) ;
  • Oh Man-Jin (Department of Food Science and Technology, Chungnam National University)
  • Published : 2005.06.01
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Abstract

This study was carried out to investigate the cathepsin B inhibition effect by Achyranthes japonica N. root extract in vitro. The methanol/$H_{2}O$(4:1, v/v) extract was fractionated by ethyl acetate(F1), chloroform(F2), chloroform/methanol(3:1, v/v)(F3) and methanol(F4). The yield of F4 in Achyranthes japonica N. root was $8.27\%$. As an index material of Achyranthes japonica N. root, 20-hydroxy ecdysone was detected by TLC, and HPLC and it's content was $0.33\%$. Three isolates(F1, F3, F4) showed the cathepsin B inhibition activity, and F4 showed the highest inhibition activity among them. In the inhibition activity on cathepsin B, leupeptin, 20-hydroxy ecdysone and F4(at the same concentration of 20-hydroxy ecdysone.) were 92, 88 and $97\%$ on BANA($N{\alpha}$-benzoyl-DL-arginine ${\beta}$-naphthylamide) substrate, and 62, 36 and $67\%$ on CLN($N{\alpha}$-CBZ(carbobenzlyoxy)-L-lysine p-nitrophenyl ester HCI) substrate, respectively.

우슬이 민간요법으로 관절염 치료에 우수한 한약재로 알려진 바 우슬 추출물이 cathepsin B 에 대한 저해력을 검토하기 위하여 각종 용매로 우슬 추출물을 순차 분획하고 분획 추출물에 대하여 cathepsin B에 대한 저해활성을 검토하였으며 우슬의 지표물질인 20-hydroxy ecdysone이 추출분획에 검출되는지 TLC 및 HPLC를 이용하여 분석하였으며 우슬의 지표물질이 cathepsin B에 대한 저해활성여부를 확인한바 다음과 같은 결과를 얻었다. 우슬 뿌리에 대해 methanol/water(4:1, v/v) 추출물을 ethyl acetate, chloroform, chloroform/methanol(3:1, v/v), methanol의 각 용매로 분획한 결과, 분획 F4(methanol 분획)에서 수율이 가장 높아 $8.27\%$를 나타내었다. 각 분획별에 따라 cathepsin B에 대한 저해활성을 측정한 결과 F4분획물에서 가장 저해활성이 높았으며 F1분획에서도 높은 저해활성을 나타내었다. F4분획물 중 우슬의 지표물질인 20-hydroxy ecdysone이 검출되었으며 우슬에 함유되어있는 함량은 $0.33\%$이었다. 20-hydroxy ecdysone에 대하여 cathepsin B저해활성을 보기 위하여 cathepsin B저해활성제로 알려진 leupeptin과 활성을 비교해본 결과 기질을 BANA로 사용하였을 경우 leupeptin은 저해율이 $92\%$인데 비하여 20- hydroxy ecdysone은 $88\%$이었으며 F4분획물은 $97\%$를 나타내었고 기질을 CLN으로 사용하였을 경우 leupeptin은 저해율이 $62\%$인데 비하여 20-hydroxy ecdysone은 $36\%$이었으며 F4추출물은 $67\%$를 나타내었다.

Keywords

References

  1. 육창수 (1990) 원색한국약용식물도감. 아카데미서적. p.175
  2. Hahn. D.R. and Lee, M.W. (1991) Studies on the constituents of Achranthis Radix(I) oleanolic acid bisdesmoside from the roots. Yakhak Hoeji, 35, 457-460
  3. Ogawa, S.N., Nishimoto, N. Okamoto and Takemoto, T. (1971) Studies on the constituents of Achyranthis Radix(VIII). The insect-moulting substances in Achyranthes genus. Yakugaku Zasshi, 91, 916-920
  4. Sun, S.P., Li, K.H. and Sun, S.S. (1985) Pharmacological studies on Achyranthes bidentata. Hanan Trad Chin. Med., 47, 39-40
  5. 이승택, 채영암 (1996) 약용작물재배. 향문사. 서울, p.159-1625
  6. 박인현, 이상래, 이상득, 송원섭 (1993) 약용식물재배. 선진문화사 서울, p.281-283
  7. Entou, Y.M. (1997) 牛膝 および 牛膝 關聯 植物の 成分硏究. 昭和大碩土論文, p.1-44
  8. De Duve, C. (1969) In 'Lyosomes in Biology and Pathology' J. T. Dingle and H. B. Fell, eds, North-Holland Pulb., Amsterdam. 1, p.3-42
  9. Ebert, W., Knoch, H., Werle, B., TRefz, G., Muley, T. and Spiess, E. (1994) Prognostic value of increased lung tumor tissue cathepsin B. Anticancer Res., 14, 895-900
  10. Lenny, J.F. (1980) Inhibitors associated with the proteinases of mammalian cells and tissues. Curr. Top. Cell. Regul., 17, 25-57
  11. Sloane B.F., Moin. K., Krepela, E. and Rozhin, J. (1990) Cathepsin B and its endogenous inhibitors:role in tumor malignancy. Cancer Metastasis Rev., 9, 333-352 https://doi.org/10.1007/BF00049523
  12. Duffy, M. J. (1996) Protease as Prognostic markers in cancer. Clin. Cancer Res., 2, 613-618
  13. Han, K.H. and Kim. S.D.(1997) Selection and identification of a strain KT-10 producing the cathepsin B inhibitor. J. Microbiol. Biotechnol., 7, 333-340
  14. Barret, A.J., Davies, M.E. and Grubb, A. (1984) The place of human ganuna-trace(cystatin C) amongst the cysteine proteinase inhibitors. Biochem. Biophysical Res. Commun., 120, 631-636 https://doi.org/10.1016/0006-291X(84)91302-0
  15. Engh. R.A., Dieckmann, W.T., Bode, E., Auerswald, A V., Huber, R. Turk and Oschkinat, H. (1993) Conformational variability of chicken cystatin. Comparison of structures determined by X-ray diffraction and NMR spectroscopy. J. Mol. Biol., 234, 1060-1069 https://doi.org/10.1006/jmbi.1993.1659
  16. Aoyagi, T., Takeuchi, T., Matsuzaki, T., Kawamura, K., Kondo, S., Hamada, M., Maeda, K. and Umezawa, H. (1969) Leupeptins, new protease inhibitor isolated from Actinomycetes. J. Antibiot, 22, 283-286 https://doi.org/10.7164/antibiotics.22.283
  17. Suda, H., Aoyagi, T, Hamada, M., Takeuchi, T. and Umezawa, H. (1972) Antipain, new protease inhibitor isolated from Actznomycetes. J. Antibiot., 25, 263-266 https://doi.org/10.7164/antibiotics.25.263
  18. Sreedharan, S.K., Verma, C.L., Caves, S.D. and Brockkleburst, S.M (1966) Demonstration that l-trans-epoxysuccinyl-L-leucylamido-(4-guanidino)butane(E-64) is one of the most effective low inhibitors of trypsin-catalysed hydrolysis. Characterization by kinetic analysis and by energy minimization and molecular dynamics simulation of the E-64-$\beta$-trypsin complex. J. Biochem., 316, 777-786
  19. Han, K.H. and Kim. S.D. (2001) Isolation and characterization of cathepsin B inhibitor produced by Streptomycetes leuteognseus KT-10. Korean J. Appl. Microbiol. Biotechnol., 29, 84-89
  20. A.O.A.C. (1995) Official Method of Analysis(16th Edition) Association of Official Analytical Chemist, Washington, D.C
  21. Perkin-Elmer Corporation. (1968) Analytical Methods for Atomic Absorption Spectroscopy, Perkin-Elmer Corp. Norwalk, Comm
  22. 김찬호, 김만욱, 최강주, 손현주, 고성룡, 김석창, 위재준, 허정남 (1991) 인삼성분분석법. 한국인삼연초연구소, 13-18
  23. Bajkowski, A.S. and Frankfater, A. (1975) Specific spectrophotometric assays for cathepsin B. Anal. Biochem., 68, 119-127 https://doi.org/10.1016/0003-2697(75)90685-5
  24. Son, K.H., Hwang, J.H., Lee, S.H., Park, J.H., Kang, S.J., Chang, S.Y. and Lee, K.S. (1999) Isolation and Quantitative determination of 20-Hydroxyecdysone from Achyranthes radix. Korean J. Pharmacogn., 30, 335-339
  25. Kim, M.S. (2003) A study on optimal cultivation conditions of medicinal components in Native Achyranthes japonica N. Ph. D. Thesis. Chonnam National University, p.21-24